A new device to mimic intermittent hypoxia in mice

Abstract

Intermittent hypoxia (hypoxia-reoxygenation) is often associated with cardiovascular morbidity and mortality. We describe a new device which can be used to submit cohorts of mice to controlled and standardised hypoxia-normoxia cycles at an individual level. Mice were placed in individual compartments to which similar gas flow parameters were provided using an open loop strategy. Evaluations made using computational fluid dynamics were confirmed by studying changes in haemoglobin oxygen saturation in vivo. We also modified the parameters of the system and demonstrated its ability to generate different severities of cyclic hypoxemia very precisely, even with very high frequency cycles of hypoxia-reoxygenation. The importance of the parameters on reoxygenation was shown. This device will allow investigators to assess the effects of hypoxia-reoxygenation on different pathological conditions, such as obstructive sleep apnoea or chronic obstructive pulmonary disease.

Figure 1. A device providing reliable and homogenous gas concentrations in individual mouse rooms.

A: Technical scheme of the circuitry 1) air compressor and tank, 2) nitrogen compressor and tank, 3) T-connectors, 4) flowmeter with an analogue valve, 5) solenoid valve, 6) check valve, 7) Y-connectors, 8) pressure regulator, 9) static mixer, 10) mouse cage, 11) mouse, 12) oxygen sensor, 13) outlet of gas mixes, 14) controller with touch panel, 15) device to measure arterial blood oxygen saturation, 16) notebook to control, visualize and save the data. Solid line: pipes, dashed line: electrical wires. B: Insertion of splitters in the airflow allows gas concentrations to be homogenised within the mouse chambers: CFD simulation of the gas flow in the mouse box without (a) and with (c) splitters; velocity profiles inside each box without (b) and with (d) splitters (section cut).

Figure 2. Control of the DC and analogue valves as a function of the required FIO_2.

The traces illustrate the strategy used when the required FIO₂ was between 21%-13% (here 18%, A) or 12-0% (here 10%, B). The first graph corresponds to traces of expected and measured gas concentrations, and the others to the operating signals controlling the valves.

Figure 3. Oxygen saturation of haemoglobin as a function of FIO₂.

Values of haemoglobin oxygen saturation are measured after 3 minutes of exposure to a FIO2 and expressed as means ± SD for the different FIO₂ values. Comparisons were made using a one-way ANOVA followed by Newman-Keuls test. *p<0.05.

Figure 4. Evolution of haemoglobin oxygen saturation in response to a cyclic change in FIO₂.

Traces from a representative animal. Saturation was assessed during four consecutive cycles of 60 s during which the hypoxic phase was progressively lengthened (15, 20, 30 and 40 s). The red line corresponds to FIO₂ in the inspired air, the black line to the transcutaneous measurement of haemoglobin oxygen saturation of the animal. Latency (1); time to peak (2) and amplitude of change in saturation (3).

Figure 5. Representative records of haemoglobin oxygen saturation during hypoxia–reoxygenation with different durations of the hypoxic phase.

During the four consecutive 60 s cycles, the hypoxic phase was progressively lengthened (15, 20, 30 and 40 s). The black line corresponds to the oxygen saturation of haemoglobin, the red line to FIO₂ in the inspired air. The three graphs represent the cycles with different FIO₂ values during the hypoxic phase (A: 12%; B: 9%; C: 6%).

Figure 6. Representative records of haemoglobin oxygen saturation at different positions in the chamber.

The black line corresponds to box 1, the green line to box 3 and the red line to box 5. FIO₂: 12% alternating with 21% every 30 s.

Figure 7. Representative records of haemoglobin oxygen saturation during chronic hypoxia-reoxygenation.

Cycles alternated 30 s of normoxia and hypoxia. The black line corresponds to a hypoxic FIO₂ of 12%, the green line to a FIO2 of 9 % and the red line to an FIO₂ of 6%.

Figure 8. Representative records of haemoglobin oxygen saturation during chronic hypoxia-reoxygenation.

Cycles alternated 15 s of hypoxia – 45 s of normoxia. The black line corresponds to a hypoxic FIO₂ of 12%, the green line to a FIO₂ of 9% and the red line to an FIO₂ of 6%.